Single-stranded (ssDNA) DNA binding proteins (SSBs) bind preferentially ssDNA in stoichiometric quantities with respect to their substrate, displaying little sequence preference and no associated ATPase activity (11). The binding is typically cooperative, though the level of cooperativity varies widely. Much effort has been spent on elucidation of the structural mechanism accounting for the cooperativity and its functional implications in the case of the filamentous phage GVP (6, 68), the phage T4 gp32 (10, 33), the adenovirus DNA binding protein (DBP) (34), the Escherichia coli SSB (19,42), and the eukaryotic replication protein A (30, 31), while little is known about the SSBs of the Herpesviridae.The herpes simplex virus type 1 (HSV-1) SSB, infected cell polypeptide 8 (ICP8), is a 128-kDa nuclear zinc metalloprotein encoded by the UL29 gene (22). By virtue of its ability to bind oligonucleotide as well as to mediate specific protein-protein interactions, it was shown to have essential functions in DNA metabolism during the viral lytic cycle. ICP8 is one of the seven ␤, or delayed-early, genes required for viral genome replication, which proceeds via a rolling circle mechanism (63, 65). Replication occurs in globular nuclear domains termed replication compartments (55) whose location is defined by the preexisting host cell nuclear architecture, most probably at the periphery of the nuclear matrix-associated ND10 domains, where the viral transactivator ICP0 and the viral input genome migrate in the early stages of infection (43, 47). Evidence has been provided that the seven essential proteins, the origin binding protein (OBP), the polymerase with its processivity factor (UL30 and UL42), the trimeric primase-helicase complex (UL52, UL5, and UL8), and ICP8, accumulate in punctuate prereplicative sites for the assembly of the multiprotein complex, or primosome, which promotes efficient genomic replication (40,71,76). During initiation, ICP8 associates with the carboxy-terminal domain of the OBP at the origin of replication to assist the ATP-dependent bidirectional origin unwinding (3,36,37,45). It enhances the polymerase processivity (29, 50, 61) and stimulates the helicase-primase activity via interaction with the UL8 subunit (4,17,21). In accord with its ability to destabilize DNA helices (5) and promote Mg-dependent complementary-strand renaturation (16), ICP8 can catalyze homologous pairing and strand transfer (7), and hence it participates in the frequent DNA recombination events. Genetic evidence implies a role for ICP8 in the regulation of viral gene expression (12,26), in agreement with the reported ICP8 affinity for polyriboadenylate (61).The DNA binding properties of ICP8 have been extensively investigated. ICP8 binds ssDNA rapidly and cooperatively, with a modest preference for the HSV genomic GC-rich sequences, holding the nucleotide filaments in an extended conformation (35,58). Optimal binding occurs at neutral pH and 150 mM salt concentration (61). Based on filter binding assays (58), nuclease prote...

Cl- efflux into various incubation media (PSS) was studied in pieces of rabbit aortae loaded with 36Cl. Replacement of HCO3-/CO2 by HEPES/O2 in the PSS increased the rate of Cl- efflux by a factor of 2.4. This effect was suppressed in Cl(-)-free PSS containing isethionate, propionate, or benzenesulfonate, but not in NO3(-)-PSS, or Br(-)-PSS. The stimulant effect of HCO3- withdrawal on Cl-efflux was reduced by 140 microM DIDS, but not by 1 mM furosemide. The Cl- efflux was temperature-dependent (Q10 = 2.3-2.5), and it was not affected on depolarisation by high [K+]o. The [Cl-]i of rabbit aorta determined by uptake studies with 36Cl, decreased slightly (by 15%) below controls in PSS containing 140 microM DIDS, but drastically (from 32.6 to 13.5 mM, i.e. by 59%) in PSS containing 1 mM furosemide. Withdrawal of HCO3-/CO2 depolarized rabbit pulmonary artery in standard PSS and in Br(-)-PSS or NO3(-)-PSS, but not in benzenesulfonate-PSS. The pHi of rabbit aorta determined by the distribution of (14C)-DMO, increased in Cl(-)-free PSS containing isethionate or glucuronate. It is concluded that transport mechanisms play a major role in the distribution of Cl- in vascular smooth muscle, and that a membrane anion carrier operates in this tissue which can transport Cl- and HCO3- across the cell membrane. This mechanism seems to be involved in the regulation of pHi. However, the known high [Cl-]i of vascular smooth muscle is rather mediated by the furosemide-sensitive Na-K-Cl cotransport than by this anion carrier.

Renal arteries of the rabbit were chemically skinned using Triton X-100. In EGTA-buffered solutions containing calmodulin and ATP, small strips of the skinned preparations were found to develop contractile force which was dependent on the concentrations of either free calcium or of free barium. However, a 220 times greater concentration of barium than of calcium was necessary for comparable effects. Quantitatively, the response to barium was dependent on the concentration of calmodulin added to the test solutions. The contractile effect of barium was partly antagonized by the calmodulin antagonist, trifluoperazine.

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